Recently, a risk of zoonotic infectious disease such as BSE (Bovine Spongiform Encephalopathy) infection and Avian Influenza virus infection, has started to be a concern. Moreover, products using ingredients or materials primarily originated from animals (such as vaccines, blood preparations, cell culture/genetic recombinant preparations, and cellular tissue medical products) have problems of a high risk where an infectious agent is mixed thereinto, and an undeniable likelihood of containing an unknown infectious agent, as well as a limitation in the inactivation treatment of infectious agents, and the like. As a countermeasure against such problems, legal countermeasures for safety have been enhanced, such as newly providing a framework for “bio-originated products” by the amendment of Japanese Pharmaceutical Law in 2003, and there has become a desire for the development of medical products free from animal-origin components.
Many attempts have been made for obtaining a culture condition not requiring a serum, such as bringing serum-free mediums free from animal-origin components into the market by respective medium manufacturers. However, in a culture method using a serum-free medium and a microcarrier where the surface of the carrier is electrically charged at an appropriate amount so as to adhere cells, as a support for culturing adhesive cells, there has been a problem in that the attachment rate to the microcarrier is reduced, making it difficult to efficiently culture a large amount of cells. As a result, conventionally, under such a culture condition, there has been used a carrier containing animal-origin components such as a microcarrier coated with denatured pig collagen, as described for example in Non Patent Document 1.
Moreover, there is disclosed in Patent Document 1, a bead for culturing animal cells which has a high cell-adhering property and a high cell-proliferating property, and, even in a serum-free culture medium, gives a cell-adhering property and a cell-proliferating property which are equivalent or more to those in serum-containing medium. However, although Patent Document 1 shows that cells can be efficiently cultured under a serum-free condition in a cell culture, there is no disclosure of a condition for producing a virus such as a virus inoculation or proliferation method.
Moreover, there is disclosed in Patent Document 2, a method of producing a virus including steps of: obtaining a vertebrate cell culture such as Vero cells; proliferating the cells only in a protein-free medium (free from serum or non-serum protein); infecting this culture with a virus; incubating the virus-infected cell culture; proliferating the virus in the medium; and producing the virus-containing medium. Furthermore, there is disclosed a usage of a protease originated from a procaryote supply source as a substance which enhances the virus activity. Patent Document 2 describes that, according to this method, the obtained virus do not contain various impurity compounds originated from a human or animal supply source, nor a protein serving as a pathogenic substance. However, in an Example a trypsin which is an animal-origin component is used as a substance which enhances the virus activity.
Furthermore, there is disclosed in Patent Document 3, a method of producing a virus infected insect cell not using a naturally-originated protein but using a cell-adhesive support having a high cell-adhering property. This production method is a method of producing a virus infected insect cell, comprising steps of using a cell-adhesive artificial peptide and/or a cell-adhesive auxiliary artificial peptide to adhere a poikilothermic animal-origin cell and a substrate, and using this cell-adhered substrate for culturing cells. Patent Document 3 describes that, by not using a naturally-originated protein for a substrate, there is no risk of containing an infectious substance such as a human-infective virus, and the safety is high. However, there is no disclosure of a cell dispersing agent free from animal-origin components or an adhesive cell originated from a homoiothermic animal. On the other hand, for the subculture of cells in Non Patent Document 1, an animal-origin protease (such as pig-origin trypsin) is used as a cell dispersing agent.    Patent Document 1: Japanese Unexamined Patent Publication No. 2003-189848    Patent Document 2: Japanese Patent Publication No. 3158157 (WO96115231 pamphlet)    Patent Document 3: Japanese Unexamined Patent Publication No. 2003-210166    Non Patent Document 1: Otfried Kistner et al. “Development of a Novel Mammalian Cell (Vero) Derived influenza Vaccine” Poster Presented at: Options for Control Of Influenza V, Okinawa, Japan, Oct. 7-11, 2003